A bifunctional dihydrofolate synthetase-folylpolyglutamate synthetase in Plasmodium falciparum identified by functional complementation in yeast and bacteria

Folate metabolism in the human malaria parasite Plasmodium falciparum is an essential activity for cell growth and replication, and the target of an i mportant class of therapeutic agents in widespread use. However, resistance to antifolate drugs is a major health problem in the developing world. To date, only two activities in this complex pathway have been targeted by ant imalarials. To more fully understand the mechanisms of antifolate resistanc e and to identify promising targets for new chemotherapies, we have cloned genes encoding as yet uncharacterised enzymes in this pathway. By means of complementation experiments using 1-carbon metabolism mutants of both Esche richia coli and Saccharomyces cerevisiae, we demonstrate here that one of t hese parasite genes encodes both dihydrofolate synthetase (DHFS) and folylp olyglutamate synthetase (FPGS) activities, which catalyse the synthesis and polyglutamation of Folate derivatives, respectively. The malaria parasite is the first known example of a eukaryote encoding both DHFS and FPGS activ ities in a single gene. DNA sequencing of this gene in antifolate-resistant strains of P. falciparum, as well as drug-inhibition assays performed on y east and bacteria expressing PfDHFS-FPGS, indicate that current antifolate regimes do not target this enzyme. As PfDHFS-FPGS harbours two activities c ritical to folate metabolism, one of which has no human counterpart, this g ene product offers a novel chemotherapeutic target with the potential to de liver a powerful blockage to parasite growth. (C) 2001 Elsevier Science B.V . All rights reserved.